ABSTRACT
PURPOSE: Congenital stromal corneal dystrophy (CSCD) is an autosomal dominant condition with clouding of the cornea due to stromal opacities. It is caused by mutations in the decorin gene (DCN) leading to the expression of a truncated form of decorin. In an attempt to replicate this condition in mice, a knock-in mouse strain, 952delT Dcn, was created. METHODS: Mice were constructed by targeted mutation. Sequencing of genomic DNA confirmed correct genotype. Mouse and human corneas, including corneas from patients with CSCD, and primary keratocyte cultures were subjected to Western blot analysis, transmission electron microscopy, and immunofluorescence microscopy. RESULTS: Histologically, the mice did not show any organ pathology. Corneas were clear, and the electron-lucent deposits observed in CSCD were not present. Furthermore, while nearly equivalent amounts of normal and truncated decorin are present in CSCD corneas, truncated decorin was hardly detectable in the mouse corneas. By immunofluorescence analysis of corneas from 952delT Dcn homozygous mice, decorin was found only in keratocytes. In primary cultures of mouse corneal explants, truncated decorin was retained intracellularly in contrast with human corneal explants where truncated decorin was exported into the culture medium. Immunofluorescence analysis revealed that native mouse decorin localized to the Golgi complex, whereas the truncated decorin accumulated in the endoplasmic reticulum (ER). CONCLUSIONS: The ER retention of truncated decorin may explain why the mouse corneas remained clear. The consequences of the decorin mutation are different in mice and humans, and 952delT Dcn knock-in mice are therefore not a suitable model for CSCD.
Subject(s)
Corneal Dystrophies, Hereditary/metabolism , Decorin/metabolism , Extracellular Space/metabolism , Animals , Cells, Cultured , Cornea/metabolism , Cornea/ultrastructure , Corneal Dystrophies, Hereditary/genetics , Corneal Dystrophies, Hereditary/pathology , Corneal Keratocytes/metabolism , Decorin/genetics , Disease Models, Animal , Endoplasmic Reticulum/metabolism , Humans , Mice, Inbred C57BL , Mice, Mutant Strains , Microscopy, Electron , MutationABSTRACT
During a period of 1.5 months, a newly established pig herd experienced a high number of mummifications and stillbirths, a high neonatal mortality rate, and many piglets with congenital tremors or hind leg ataxia. After clinical and histological investigations, the submitted animals were divided into 4 groups: mummified or stillborn (N = 6), live born with myocarditis (N = 5) (average age 22.8 days), live born without myocarditis (N = 14) (average age 20.0 days), and control animals from a different herd (N = 5) (newborn). Statistically significant differences were observed in the mean porcine circovirus 2 (PCV2) load among the 4 groups in the liver (P < 0.0001). The presence of PCV2 antigen within the myocardial lesions was confirmed by immunohistochemistry. A high load of PCV2 DNA was observed in myocardium, liver, and spleen from mummified or stillborn piglets (>1 x 10(7) copies per 500 ng DNA), lower in piglets with myocarditis (>1 x 10(5) copies per 500 ng DNA), and even further lower in pigs without myocarditis (<1 x 10(5) copies per 500 ng DNA), whereas no PCV2 DNA was detected in the control animals. Myocardium, liver, and spleen were well suited for routine testing of fetuses and young piglets by quantitative real-time polymerase chain reaction. Neither porcine parvovirus nor encepaholomyocarditis virus was detected. These results indicate that the PCV2 infection might have been of etiological importance for the fetal deaths and piglet mortality observed in this herd.